Study On Transformation Of AGPase Gene In Maize

Maize is one of the most important crops in the world. Not only maize is used as forage, but also used as industrial materials. Maize starch is commonly used as industrial product, such as textiles, cosmetics, pharmaceuticals, biofuel, biodegradable plastic products, industrial special materials. With the increasing of requirements in modern industry, breeding new high starch maize is the key to solve the problem.In the metabolism of maize endosperm starch synthesis,AGPase catalyzes Glu-1-phosphate and ATP to form the activated glucosyl donor ADPG and pyrophosphate. Starch synthases (SS) catalyzes ADPG to elongate linear chains by forming a new ?-(1→4) linkages. The linear chains construct the base structure of starch. AGPase is the key enzyme in the maize endosperm starch synthesis. The study of AGPase is mainly in mechanism research. There is few report about the application research of AGPase. In this study, in order to increasing the expression of AGPase and starch in maize endosperm, the genes of large and small subunit of maize and arabidopsis AGPase were cloned and transformed into inbred lines by transgenic technology. The main results were as follow:(1) The stable genetic transformation system need a good recipient system. Using immature embryo which was from 42 excellent maize inbred lines as explant, induce embryogenic callus and differentiate regeneration plantlets. Five genotypes could induce embryogenic callus after selection. Those explants, from plumule, immature embryo, germ, shoot, were also obtained regeneration plantlets by organogenesis pathway. This study established four stable genetic transformation recipient system.(2) Four genes were cloned, which were control the synthesis of large and small subunit in maize and arabidopsis AGPase, and the rice endosperm specific promoter RP5. Those efficient plant expression vectors, pRP5-ZmAGPL, pRP5-ZmAGPS and pRP5-AtAGPL were successfully constructed.(3) Constructed plant expression vectors were transformed into embryogenic callus, immature embryo and shoot, form the excellent inbred lines by Agrobacterium tumefaciens mediated. Three genetic transformation system were established. Transformation ratio was 5%, 4.4% and 12.5%.(4) Through pollen-tube pathway, the AGPase plant expression vectors were transformed into 15 maize inbred lines, with applying single gene transformation and polygene co-transformation. 53853 grains T0 generation seed were obtained. Some maize inbred lines also got T1 generation seed. The transformation ratio was 2.4%.This research has the important theoretical and practical value for using transgenic technology to improve maize endosperm starch content in the future.